ZoBell 



48 



Marine Microbiology 



Within the limits of the experimental error, synthetic sea water or sea 

 water diluted with an equal volume of distilled water gave results which 

 were comparable with results obtained by using natural sea water. For 

 routine work sea water diluted 3 :i with distilled water is recommended for 

 dilution water blanks. This so-called 75 per cent sea water provides for 

 the prolonged survival of marine bacteria, and unlike undiluted sea water, 

 no precipitate develops in it after sterilization and standing in soft glass 

 receptacles. 



It is noteworthy that marine bacteria are equally sensitive to both 

 hypertonicity and to hypotonicity as indicated by the results with sea 

 water to which 3 per cent sodium chloride had been added. Working with 

 " fresh- water " organisms. Tanner and Houston (1940) concluded that 

 bacteria are quite resistant to changes in osmotic pressure. Our observa- 

 tions confirm those of Korinek (1926) that " fresh- water " bacteria are 

 more resistant to changes in osmotic pressure than are marine bacteria. 

 Additional information on this subject is given in Chapter VIII. 



The successive dilution method for enumerating bacteria : — The suc- 

 cessive dilution method, also called the extinction or minimum dilution 

 method, consists of diluting the material to be examined usually by pow- 

 ers of 10, and inoculating equal volumes of the diluted material into liquid 

 media (Halvorson and Ziegler, 1933a). The reciprocal of the highest 

 dilution which yields positive results is regarded as the most probable 

 number of bacteria present. Thus the occurrence of growth from the in- 

 oculation of i.o ml. of a 1:1000 dilution but not of a 1:10,000 dilution 

 indicates that the original material contained at least 1000 viable bacteria 

 per ml. 



As pointed out by Halvorson and Ziegler (19336), the accuracy of 

 the dilution method depends primarily upon the uniform distribution of 

 cells in the solution, the number of dilutions used, the number of tubes 

 inoculated with each dilution, and the fitness of the media to support the 

 growth of the bacteria. Inoculating each of ten tubes with three effective 

 dilutions, ZoBell (1941a) analyzed 38 samples of marine mud and sea 

 water by the successive dilution method and also by duplicate plate 

 counts. Representative results are given in Table XII. The nutrient 

 composition of the media was identical. 



Table XII. — Most probable number of bacteria in samples of marine materials indicated 

 by the successive dilution method and plate counts: — 



There was a fair agreement between the results obtained by the suc- 

 cessive dilution method and plating procedures, the former being an aver- 

 age of 12 per cent higher than plate counts. Similarly, Butterfield 

 (19336) found that if enough tubes were used under standardized condi- 



